Microbiology (medical)

‘Candidatus Liberibacter asiaticus’ is known as the most pathogenic organism associated with citrus greening disease. Since its publicized emergence in Florida in 2005, ‘Ca. L. asiaticus’ remains unculturable. Currently, a limited number of potential disease effectors have been identified through in silico analysis. Therefore, these potential effectors remain poorly characterized and do not fully explain the complexity of symptoms observed in citrus trees infected with ‘Ca. L. asiaticus.’ LotP has been identified as a potential effector and have been partially characterized. This protein retains structural homology to the substrate binding domain of the Lon protease. LotP interacts with chaperones like GroEL, Hsp40, DnaJ, and ClpX and may exercise its biological role through interactions with different proteins involved in proteostasis networks. Here, we evaluate the interactome of LotP—revealing a new protein–protein interaction target (Lon-serine protease) and its effect on citrus plant tissue integrity. We found that via protein–protein interactions, LotP can enhance Lon protease activity, increasing the degradation rate of its specific targets. Infiltration of purified LotP strained citrus plant tissue causing photoinhibition and chlorosis after several days. Proteomics analysis of LotP tissues recovering after the infiltration revealed a large abundance of plant proteins associated with the stabilization and processing of mRNA transcripts, a subset of important transcription factors; and pathways associated with innate plant defense were highly expressed. Furthermore, interactions and substrate binding module of LotP suggest potential interactions with plant proteins, most likely proteases.

Magnetotactic bacteria (MTB) are a group of microbes that biomineralize membrane-bound, nanosized magnetite (Fe3O4), and/or greigite (Fe3S4) crystals in intracellular magnetic organelle magnetosomes. MTB belonging to the Nitrospirae phylum can form up to several hundreds of Fe3O4 magnetosome crystals and dozens of sulfur globules in a single cell. These MTB are widespread in aquatic environments and sometimes account for a significant proportion of microbial biomass near the oxycline, linking these lineages to the key steps of global iron and sulfur cycling. Despite their ecological and biogeochemical importance, our understanding of the diversity and ecophysiology of magnetotactic Nitrospirae is still very limited because this group of MTB remains unculturable. Here, we identify and characterize two previously unknown MTB populations within the Nitrospirae phylum through a combination of 16S rRNA gene-based and genome-resolved metagenomic analyses. These two MTB populations represent distinct morphotypes (rod-shaped and coccoid, designated as XYR, and XYC, respectively), and both form more than 100 bullet-shaped magnetosomal crystals per cell. High-quality draft genomes of XYR and XYC have been reconstructed, and they represent a novel species and a novel genus, respectively, according to their average amino-acid identity values with respect to available genomes. Accordingly, the names Candidatus Magnetobacterium cryptolimnobacter and Candidatus Magnetomicrobium cryptolimnococcus for XYR and XYC, respectively, were proposed. Further comparative genomic analyses of XYR, XYC, and previously reported magnetotactic Nitrospirae reveal the general metabolic potential of this MTB group in distinct microenvironments, including CO2 fixation, dissimilatory sulfate reduction, sulfide oxidation, nitrogen fixation, or denitrification processes. A remarkably conserved magnetosome gene cluster has been identified across Nitrospirae MTB genomes, indicating its putative important adaptive roles in these bacteria. Taken together, the present study provides novel insights into the phylogenomic diversity and ecophysiology of this intriguing, yet poorly understood MTB group.

The Asian citrus psyllid, Diaphorina citri, is the vector of the bacterium “Candidatus Liberibacter asiaticus” (Las), associated with the devastating, worldwide citrus disease huanglongbing. In order to explore the molecular interactions of this bacterium with D. citri during the vector acquisition process, cDNA libraries were sequenced on an Illumina platform, obtained from the gut of adult psyllids confined in healthy (H) and in Las-infected young shoots (Las) for different periods of times (I = 1/2 days, II = 3/4 days, and III = 5/6 days). In each sampling time, three biological replicates were collected, containing 100 guts each, totaling 18 libraries depleted in ribosomal RNA. Reads were quality-filtered and mapped against the Chinese JXGC Las strain and the Floridian strain UF506 for the analysis of the activity of Las genome and SC1, SC2, and type 3 (P-JXGC-3) prophages of the studied Las strain. Gene activity was considered only if reads of at least two replicates for each acquisition access period mapped against the selected genomes, which resulted in coverages of 44.4, 79.9, and 94.5% of the JXGC predicted coding sequences in Las I, Las II, and Las III, respectively. These genes indicate an active metabolism and increased expression according to the feeding time in the following functional categories: energy production, amino acid metabolism, signal translation, cell wall, and replication and repair of genetic material. Pilins were among the most highly expressed genes regardless of the acquisition time, while only a few genes from cluster I of flagella were not expressed. Furthermore, the prophage region had a greater coverage of reads for SC1 and P-JXGC-3 prophages and low coverage in SC2 and no indication of activity for the lysis cycle. This research presents the first descriptive analysis of Las transcriptome in the initial steps of the D. citri gut colonization, where 95% of Las genes were active.

Anoxygenic phototrophic bacteria can be important primary producers in some meromictic lakes. Green sulfur bacteria (GSB) have been detected in ferruginous lakes, with some evidence that they are photosynthesizing using Fe(II) as an electron donor (i.e., photoferrotrophy). However, some photoferrotrophic GSB can also utilize reduced sulfur compounds, complicating the interpretation of Fe-dependent photosynthetic primary productivity. An enrichment (BLA1) from meromictic ferruginous Brownie Lake, Minnesota, United States, contains an Fe(II)-oxidizing GSB and a metabolically flexible putative Fe(III)-reducing anaerobe. “Candidatus Chlorobium masyuteum” grows photoautotrophically with Fe(II) and possesses the putative Fe(II) oxidase-encoding cyc2 gene also known from oxygen-dependent Fe(II)-oxidizing bacteria. It lacks genes for oxidation of reduced sulfur compounds. Its genome encodes for hydrogenases and a reverse TCA cycle that may allow it to utilize H2 and acetate as electron donors, an inference supported by the abundance of this organism when the enrichment was supplied by these substrates and light. The anaerobe “Candidatus Pseudopelobacter ferreus” is in low abundance (∼1%) in BLA1 and is a putative Fe(III)-reducing bacterium from the Geobacterales ord. nov. While “Ca. C. masyuteum” is closely related to the photoferrotrophs C. ferroooxidans strain KoFox and C. phaeoferrooxidans strain KB01, it is unique at the genomic level. The main light-harvesting molecule was identified as bacteriochlorophyll c with accessory carotenoids of the chlorobactene series. BLA1 optimally oxidizes Fe(II) at a pH of 6.8, and the rate of Fe(II) oxidation was 0.63 ± 0.069 mmol day–1, comparable to other photoferrotrophic GSB cultures or enrichments. Investigation of BLA1 expands the genetic basis for phototrophic Fe(II) oxidation by GSB and highlights the role these organisms may play in Fe(II) oxidation and carbon cycling in ferruginous lakes.

Bactericera cockerelli es una plaga de importancia económica en solanáceas en México, por los amarillamientos que causa en los cultivos, así como por la transmisión de Candidatus Liberibacter solanacearum. Se describen variantes genéticas de este insecto, las cuales se relacionan con su capacidad para fungir como vector. En México, la distribución de B. cockerelli es muy amplia y se carece de información acerca de sus características morfológicas y genéticas. El objetivo de esta investigación fue caracterizar morfológica y genéticamente a B. cockerelli y detectar la presencia de Ca. L. solanacearum en poblaciones de B. cockerelli de las zonas productoras de solanáceas en México. Para lo cual se muestrearon 35 localidades de 13 estados, sobre cultivos de chile, tomate, berenjena y papa, bajo diferentes sistemas de producción. Se midieron las variables largas de cuerpo (LC), largas de abdomen (LAB) y ancho de abdomen (AAB) en insectos de cada población, se detectó la presencia de Ca. L. solanacearum en los 13 estados muestreados, donde los machos presentaron el mayor porcentaje de insectos positivos. La presencia de Ca. L. solanacearum no se vio influenciada por el hospedero o el sistema de producción, sino por la presencia de B. cockerelli.

Citrus Huanglongbing (HLB; yellow shoot disease) is associated with an unculturable α-proteobacterium “Candidatus Liberibacter asiaticus” (CLas). HLB was found in southern California in 2012, and the current management strategy is based on suppression of the Asian citrus psyllid (Diaphorina citri) that transmits CLas and removal of confirmed CLas-positive trees. Little is known about Asian citrus psyllid-associated bacteria and citrus-associated bacteria in the HLB system. Such information is important in HLB management, particularly for accurate detection of CLas. Recent advancements in next-generation sequencing technology provide new opportunities to study HLB through genomic DNA sequence analyses (metagenomics). In this study, HLB-related bacteria in Asian citrus psyllid and citrus (represented by leaf midrib tissues) samples from southern California were analyzed. A metagenomic pipeline was developed to serve as a prototype for future bacteriomic research. This pipeline included steps of next-generation sequencing in Illumina platform, de novo assembly of Illumina reads, sequence classification using the Kaiju tool, acquisition of bacterial draft genome sequences, and taxonomic validation and diversity evaluation using average nucleotide identity. The identified bacteria in Asian citrus psyllids and citrus together included Bradyrhizobium, Buchnera, Burkholderia, “Candidatus Profftella armature,” “Candidatus Carsonella ruddii,” CLas, Mesorhizobium, Paraburkholderia, Pseudomonas, and Wolbachia. The whole genome of a CLas strain recently found in San Bernardino County was sequenced and classified into prophage typing group 1 (PTG-1), one of the five known CLas groups in California. Based on sequence similarity, Bradyrhizobium and Mesorhizobium were identified as possible source that could interfere with CLas detection using the 16S rRNA gene-based PCR commonly used for HLB diagnosis, particularly at low or zero CLas titer situation.

Understanding how phytoplasmas move and multiply within the host plant is fundamental for plant–pathogen interaction studies. In recent years, the tomato has been used as a model plant to study this type of interaction. In the present work, we investigated the distribution and multiplication dynamics of one strain of ‘Candidatus Phytoplasma (Ca. P.) solani’ (16SrXII-A) in tomato (Solanum lycopersicum L., cv. Micro-Tom) plants. We obtained infected plants by grafting, a fast and effective method to maintain phytoplasma infection. In planta spread and multiplication of ‘Ca. P. solani’ was monitored over time using qualitative and quantitative qPCR. Root, apical shoot, lower leaves, and upper leaves were sampled at each sampling time. We hypothesized that ‘Ca. P. solani’ from the grafting site reached firstly the highest leaf, the apex and the roots; subsequently, the phytoplasmas spread to the rest of the upper leaves and then progressively to the lower leaves. Significant differences were found in ‘Ca. P. solani’ titer among different plant tissues. In particular, the concentration of phytoplasma in the roots was significantly higher than that in the other plant compartments in almost all the sampling dates. Since the roots show rapid colonization and the highest concentration of phytoplasmas, they represent the ideal tissue to sample for an early, sensitive and robust diagnosis.

CandidatusMicrothrix is one of the most common bulking filamentous microorganisms found in activated sludge wastewater treatment plants (WWTPs) across the globe. One species,Ca.M. parvicella, is frequently observed, but global genus diversity, as well as important aspects of its ecology and physiology, are still unknown. Here, we use the MiDAS ecosystem-specific 16S rRNA gene database in combination with amplicon sequencing of Danish and global WWTPs to investigateCa.Microthrix spp. diversity, distribution, and factors affecting their global presence. Only two species were abundant across the world confirming low diversity of the genus: the dominantCa.M. parvicella and an unknown species typically present along withCa.M. parvicella, although usually in lower abundances. Both species were mostly found in Europe at low-to-moderate temperatures and their growth was favored in municipal WWTPs with advanced process designs. As no isolate is available for the novel species, we propose the name “CandidatusMicrothrix subdominans.” Ten high-quality metagenome-assembled genomes recovered from Danish WWTPs, including 6 representing the novelCa.M. subdominans, demonstrated high genetic similarity between the two species with a likely preference for lipids, a putative capability to reduce nitrate and nitrite, and the potential to store lipids and poly-P.Ca.M. subdominans had a potentially more versatile metabolism including additional sugar transporters, higher oxygen tolerance, and the potential to use carbon monoxide as energy source. Newly designed fluorescencein situhybridization probes revealed similar filamentous morphology for both species. Raman microspectroscopy was used to quantify thein situlevels of intracellular poly-P. Despite the observed similarities in their physiology (both by genomes andin situ), the two species showed different seasonal dynamics in Danish WWTPs through a 13-years survey, possibly indicating occupation of slightly different niches. The genomic information provides the basis for future research intoin situgene expression and regulation, while the new FISH probes provide a useful tool for further characterizationin situ. This study is an important step toward understanding the ecology ofCa.Microthrix in WWTPs, which may eventually lead to optimization of control strategies for its growth in this ecosystem.

AbstractExtracellular DNA is a major macromolecule in global element cycles, and is a particularly crucial phosphorus, nitrogen and carbon source for microorganisms in the seafloor. Nevertheless, the identities, ecophysiology and genetic features of DNA-foraging microorganisms in marine sediments are largely unknown. Here, we combined microcosm experiments, DNA stable isotope probing (SIP), single-cell SIP using nano-scale secondary isotope mass spectrometry (NanoSIMS) and genome-centric metagenomics to study microbial catabolism of DNA and its subcomponents in marine sediments.13C-DNA added to sediment microcosms was largely degraded within 10 d and mineralized to13CO2. SIP probing of DNA revealed diverse ‘CandidatusIzemoplasma’,Lutibacter,Shewanellaand Fusibacteraceae incorporated DNA-derived13C-carbon. NanoSIMS confirmed incorporation of13C into individual bacterial cells of Fusibacteraceae sorted from microcosms. Genomes of the13C-labelled taxa all encoded enzymatic repertoires for catabolism of DNA or subcomponents of DNA. Comparative genomics indicated that diverse ‘CandidatusIzemoplasmatales’ (former Tenericutes) are exceptional because they encode multiple (up to five) predicted extracellular nucleases and are probably specialized DNA-degraders. Analyses of additional sediment metagenomes revealed extracellular nuclease genes are prevalent among Bacteroidota at diverse sites. Together, our results reveal the identities and functional properties of microorganisms that may contribute to the key ecosystem function of degrading and recycling DNA in the seabed.